During the first two years of funding of this 3-year grant we successfully addressed one highly significant weakness of most current systems to evaluate gene therapy approaches to AIDS: the lack of a reliable and robust in vivo system that could recapitulate all aspects of human T cell neogenesis. We tested the hypothesis that human hematopoietic stem cells (HSC) capable of repopulating long-term the bone marrow of recipient immune deficient mice could retain their full hematopoietic potential and give rise to T cell progenitors. As indicated in the body of this grant, our results have fully validated this hypothesis. In our Progress Report and Preliminary Data we show the development of our novel small animal model that allows the in vivo evaluation of gene transfer into human T cell progenitors. We show that human HSC transplanted into NOD/SCID mice previously implanted with autologous human thymic/liver tissue provide long-term in vivo repopulation of human, myeloid, B and T cell compartments. Furthermore, these animals also had readily detectable levels of plasmacytoid (CD123+) and myeloid (CD11c+) dendritic cells. Analysis of the T cell compartment of implanted/transplanted mice demonstrated the presence of single positive T cells as well as both naive and memory T cells in the periphery, bone marrow and spleen. Furthermore, using lentivirus mediated gene transfer, we show that ex vivo transduced human HSC can engraft and give rise to all hematopoietic lineages while sustaining long-term transgene expression. In particular, we show transgene expression in both human thymocytes and T cells derived from ex vivo transduced human HSC. Now that we have established this system, in this competitive renewal we propose to proceed with the next logical step of our studies: the in vivo characterization of transgene expression in the human T cell compartment in these mice and the in vivo evaluation of molecular inhibitors of HIV replication.